Project Summary/Abstract: Sphingolipids are a family of membrane lipids with important structural roles in the regulation of the fluidity and subdomain structure of the lipid bilayer, especially lipid rafts. Ceramide (Cer) is the key metabolite for all sphingolipid biosynthesis. Cer and its metabolite sphingosine-1-phosphate (S1P) are signaling sphingolipids; implicated in several human diseases associated with inflammation, tumorigenesis, diabetes, and neurodegeneration. Cer signals primarily for apoptosis, whereas S1P has an opposing intracellular role in cell survival. S1P also signals paracellularly via its receptors (S1P1-5), which are mostly present in vascular endothelial cells and T cells and signals for adhesion, migration, inflammation, and neovascularization. Consistent with its role in apoptosis, ceramide has recently been shown to be involved in photoreceptor cell death. Our preliminary data show ceramide levels are increased during retinal degeneration in several models of inherited and stress-induced retinal degeneration. We also observed increased S1P levels and expression in degenerating retinas. S1P is known to have inhibitory effect on ceramide synthesis. In pilot studies, we determined that S1P is a competitive inhibitor of sphingomyelinase (SMase) also, another group of enzymes responsible for ceramide production in cells, suggesting a role for S1P in the regulated feedback of Cer production. When we injected Cer into the rat vitreous, we observed severe inflammation followed by loss of retinal function and photoreceptor cell death. We further found that FTY720, a de novo Cer synthesis inhibitor, blocked Cer production in the rat retina and protected rods from light-induced degeneration. Our preliminary results show an active sphingolipid metabolism in the retina and suggest that the delicate balance between ceramide and S1P is important in maintaining normal retinal structure and function. In this proposal we will test the hypothesis that the dysregulated balance between Cer and S1P in the retina leads to retinal inflammation and cell death. The experiments proposed in four specific aims will focus on elucidating the physiological role of Cer in retinal degenerative diseases, understanding how Cer levels are regulated, exploring the therapeutic potential of inhibitors that target Cer synthesis or the enzymes that can degrade Cer, determining how S1P is related to Cer metabolism in the retina, and how important is the balance of Cer and S1P in maintaining retinal homeostasis and function. This proposal will explore the role of Ceramide and S1P in retinal physiology and diseases. This is a novel and underserved area of retinal research and has relevance to many forms of human retinal dystrophies including age-related macular degeneration, diabetic retinopathy, and retinitis pigmentosa. Mechanistic studies will likely identify novel pathways and novel targets for therapeutic intervention.